tailieunhanh - Báo cáo khoa học: Intracellular trafficking of endogenous fibroblast growth factor-2

We have previously reported how the release of fibroblast growth factor-2 (FGF-2) is mediated by shed vesicles. In the present study, we address the question of how newly synthesized FGF-2 is targeted to the budding vesicles. | ỊFEBS Journal Intracellular trafficking of endogenous fibroblast growth factor-2 Simona Taverna Salvatrice Rigogliuso Monica Salamone and Maria Letizia Vittorelli Dipartimento di Biologia Cellulare e dello Sviluppo Universita di Palermo Italy Keywords FGF-2 microfilaments microtubules secretion of leaderless proteins shed vesicles Correspondence M. L. Vittorelli Dipartimento di Biologia Cellulare e dello Sviluppo University di Palermo Viale delle Scienze ed. 16 90128 Palermo Italy Fax 39 0 9165 77430 Tel 39 0 9165 77407 E-mail mlvitt@ Received 2 August 2007 revised 18 January 2008 accepted 31 January 2008 doi We have previously reported how the release of fibroblast growth factor-2 FGF-2 is mediated by shed vesicles. In the present study we address the question of how newly synthesized FGF-2 is targeted to the budding vesicles. Considering that in vitro cultured Sk-Hep1 hepatocarcinoma cells release FGF-2 and shed membrane vesicles only when cultured in the presence of serum we added serum to starved cells and monitored intracellular movements of the growth factor. FGF-2 was targeted both to the cell periphery and to the nucleus and nucleolus. Movements toward the cell periphery were not influenced by drugs affecting microtubules but were inhibited by cytocalasin B. Involvement of actin in FGF-2 trafficking toward the cell periphery was supported by coimmunoprecipitation and immune localization experiments. Colocalization of FGF-2 granules moving to the cell periphery and FM4-64-labelled intracellular lipids were not observed. Ouabain and methylamine two inhibitors of FGF-2 release were analyzed for their effects on FGF-2 intracellular localization and on vesicle shedding. Ouabain inhibited FGF-2 movements toward the cell periphery. The FGF-2 content of shed vesicles was therefore reduced. Methylamine inhibited vesicle shedding in its presence FGF-2 clustered at the cell periphery but the rate of its release decreased. FGF-2